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Identification of proline-rich protein 11 as a major regulator in mouse spermatogonia maintenance via an increase in BMI1 protein stability

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Abstract

Background

Spermatogenesis accompanied by self-renewal and differentiation of spermatogonia under complicated regulation is crucial for male fertility. Our previous study demonstrated that the loss of the B-lymphoma Mo-MLV insertion region 1 (BMI1) could cause male infertility and found a potential interaction between BMI1 and proline-rich protein 11 (PRR11); however, the specific co-regulatory effects of BMI1/PRR11 on spermatogonia maintenance remain unclear.

Methods and results

The expression of PRR11 was downregulated in a mouse spermatogonia cell line (GC-1) via transfection with PRR11-siRNAs, and PRR11 knockdown was verified by real-time reverse transcriptase polymerase chain reaction (RT-qPCR). The proliferative activity of GC-1 cells was determined using the cell counting kit (CCK-8), colony formation, and 5-ethynyl-2-deoxyuridine (EdU) incorporation assay. A Transwell assay was performed to evaluate the effects of PRR11 on GC-1 cell migration. A terminal deoxynucleotidyl transferase dUTP nick end labeling assay was used to measure GC-1 cell apoptosis. Furthermore, co-immunoprecipitation, RT-qPCR, and western blot analyses were used for investigating the regulatory mechanisms involved in this regulation. It was found that downregulation of PRR11 could cause a marked inhibition of proliferation and migration and induced apoptosis in GC-1 cells. Moreover, silencing of PRR11 obviously led to a reduction in the BMI1 protein level. PRR11 was found to interact with BMII at the endogenous protein level. PRR11 knockdown produced a decrease in BMI1 protein stability via an increase in BMI1 ubiquitination after which derepression in the transcription of protein tyrosine phosphatase receptor type M (Ptprm) occurred. Importantly, knockdown of Ptprm in PRR11-deficient GC-1 cells led to a reversal of proliferation and migration of GC-1 cells.

Conclusions

This study uncovered a novel mechanism by which PRR11 cooperated with BMI1 to facilitate GC-1 maintenance through targeting Ptprm. Our findings may provide a better understanding of the regulatory network in spermatogonia maintenance.

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Data availability

The data sets analyzed during the current study are available from the corresponding author upon reasonable request.

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Funding

This work was supported by the National Natural Science Foundation of China (Grant Nos.: 81901532 and 81901533); the Natural Science Foundation of Jiangsu Province (Grant No.: BK20190188); the Gusu Health Talent Program of Suzhou (Grant No.: GSWS2020068); the Suzhou Science and Technology Development Plan (Grant Nos.: SS202060 and SZM2021010); and the Introduce Project of Clinical Medicine Experts of Suzhou Industrial Park (Grant Number: SZYQTD202104).

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Author notes

  1. Jiajia Xue, Tiantian Wu and Chao Huang have contributed equally to this work.

Authors and Affiliations

  1. Suzhou Dushu Lake Hospital (Dushu Lake Hospital Affiliated to Soochow University), Suzhou, 215124, China

    Jiajia Xue, Minghua Shu & Jinxing Lv

  2. State Key Laboratory of Reproductive Medicine, Center for Reproduction and Genetics, Suzhou Municipal Hospital, The Affiliated Suzhou Hospital of Nanjing Medical University, Gusu School, Nanjing Medical University, Suzhou, 215002, China

    Tiantian Wu, Chao Huang, Cong Shen & Bo Zheng

  3. State Key Laboratory of Reproductive Medicine, Department of Histology and Embryology, Nanjing Medical University, Nanjing, 211166, China

    Tiantian Wu

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Contributions

Conceived and designed the experiments: BZ, JL; Performed the experiments: JX, YW, CH, CS; Statistical analysis: JX, MS; Wrote the paper: JX, BZ. All authors read and approved the final manuscript.

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Correspondence to Bo Zheng or Jinxing Lv.

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Xue, J., Wu, T., Huang, C. et al. Identification of proline-rich protein 11 as a major regulator in mouse spermatogonia maintenance via an increase in BMI1 protein stability. Mol Biol Rep 49, 9555–9564 (2022). https://doi.org/10.1007/s11033-022-07846-8

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